Ligand-Guided Selection and Biochemical Characterization of DNA Aptamers Against the Beta-2 Adrenergic Receptors

Author

• Agbor Otu Egbe Vydaline, CUNY Graduate CenterFollow

Date of Degree

6-2026

Document Type

Doctoral Dissertation

Degree Name

Doctor of Philosophy

Program

Biochemistry

Advisor

Prabodhika Mallikaratchy

Committee Members

Wayne Harding

Mark Emerson

Amedee Des Georges

Alex Thomsen

Subject Categories

Life Sciences

Keywords

DNA aptamers, Beta-2 adrenergic receptor, GPCRs, cell-SELEX, Ligand-guided selection, Agonists, Antagonist

Abstract

G protein-coupled receptors (GPCRs) are key to cell signaling and major targets for therapy. However, their conformational flexibility, membrane placement, and variability pose challenges for designing specific ligands. The β₂-adrenergic receptor(β₂AR), a well-characterized GPCR with important physiological and pharmacological roles, serves as an ideal model for tackling these issues. In this study, we used ligand-guided selection (LIGS) to select DNA aptamers that target the β₂AR. Aptamers are synthetic nucleic acid molecules that bind their targets with high affinity and specificity due to their unique three-dimensional structures. They are produced via systematic evolution of ligands by exponential enrichment (SELEX), but to steer enrichment toward biologically relevant conformations, we applied the LIGS method with ongoing monitoring by next-generation sequencing. Bioinformatics analysis of the enriched libraries revealed dominant sequence families, enabling us to select 20 promising aptamer candidates based on fold-enrichment ratios. These candidates were synthesized and tested using various biochemical assays and molecular docking. Using this integrated approach, we identified three lead aptamers EV β₂AR 12, 16, and 19, with nanomolar affinity, competitive and selective binding to the extracellular loops (ECL2 and ECL3) of β₂AR, mediated by non-covalent interactions including hydrogen bonds, Van der Waals, and electrostatic interactions. The three aptamers that originated from LIGS pools partitioned with the agonists isoproterenol and epinephrine mimic their functional activity, leading to receptor internalization. This work demonstrates the potential of small molecules with LIGS to target complex receptors and showcases DNA aptamers as effective molecular probes for studying β₂AR.

Recommended Citation

Egbe Vydaline, Agbor Otu, "Ligand-Guided Selection and Biochemical Characterization of DNA Aptamers Against the Beta-2 Adrenergic Receptors" (2026). CUNY Academic Works.
https://academicworks.cuny.edu/gc_etds/6637